SUMMARY: PROJECT 4 Genomic instability is a fundamental feature of human cancer. Certain cancer types harbor underlying defects in DNA repair and thus are particularly susceptible to therapeutic strategies introducing DNA damage (e.g., BRCA1 mutant breast and ovarian cancers). In prostate cancer (PCa), structural genomic rearrangements, including translocations and copy number aberrations, are a common mechanism driving tumorigenesis. However, genetic alterations in PCa predisposing to chromosomal rearrangements remain largely undefined. Whole genome sequencing demonstrates that PCa harboring recurrent point mutations in SPOP (SPOPmut) display significantly higher numbers of genomic rearrangements compared with other clinically localized PCas. These observations raise the possibility that SPOP mutations, early events in PCa tumorigenesis, lead to genomic instability. Preliminary studies demonstrate that around 10% of PCa are SPOPmut, and these represent a distinct molecular subclass. Preliminary studies in vitro demonstrate that cells expressing SPOP mutations accumulate DNA double-strand breaks (DSBs) due to altered DNA repair processes. Relevant to this SPORE project, we have shown that SPOP mutation results in increased sensitivity to DNA-damaging therapeutic agents such as ionizing radiation and poly (ADP-ribose) polymerase (PARP) inhibitors. Based on these observations, we hypothesize that SPOP mutation promotes accumulation of genomic rearrangements through impaired DSB repair and the SPOPmut subclass of PCa may be selectively responsive to DNA- damaging therapeutics, nominating alternative treatment strategies. We will test this hypothesis using a novel in vitro platform (organoids) to study endogenous SPOP mutations, in vivo mouse models testing the effect SPOP mutation on DNA damage response, and patient samples to define the response of SPOPmut patients to DNA damaging therapies. This highly collaborative project brings together established investigators supported by the PCF and SU2C- PCF who will now study the translational aspects of SPOPmut PCa. Using organoids, state-of-the-art mouse models, and cutting edge patient analyses, we will impact how clinical trials using DNA-damaging agents are rationally designed and analyzed.